Syphilis PCR

Consistent with O. Reg. 671/92 of the French Language Services Act, laboratory testing information on this page is only available in English because it is scientific or technical in nature and is for use only by qualified health care providers and not by members of the public.

Background
This page provides testing information for real-time polymerase chain reaction (PCR) testing for direct detection of Treponema pallidum subsp. pallidum (T. pallidum), the causative agent of syphilis at Public Health Ontario (PHO). For information regarding other testing options for syphilis, refer to the following PHO webpages:

*Uncontrolled print copy. Valid only on day of print: 23 June 2025.

Testing Indications

Syphilis PCR is a nucleic acid amplification test (NAAT) that is used for the direct detection of T. pallidum deoxyribonucleic acid (DNA) in skin and mucosal lesions and should be used in conjunction with syphilis serology.

Testing is recommended for individuals with suspected primary syphilis (presence of chancre/lesion, which is usually painless). Syphilis PCR may be helpful in diagnosing early primary syphilis when a lesion is present, and syphilis serology is non-reactive (i.e. prior to seroconversion).

Syphilis PCR may be considered as an adjunct diagnostic tool for lesions in the investigation of secondary syphilis. Generally, syphilis serology results, an assessment of clinical symptoms, and a review of the individual’s history and risk factors is sufficient to make a diagnosis of secondary syphilis.

PHO has not validated syphilis PCR for neonatal specimens. For the investigation of congenital syphilis, please consult a PHO Microbiologist prior to submitting specimens.

Acceptance/Rejection Criteria

Acceptable specimens: swab of lesion or lesion exudates from skin and mucosal sites placed in Universal Transport Media (UTM)

The following will be rejected:

Specimens collected using expired collection kits.
Incorrect specimens - only lesion swabs have been validated. Other specimen types (e.g. CSF and other body fluids) require PHO Microbiologist approval prior to collecting, otherwise they will be rejected.
Incorrect collection kits or dry swabs.
Mismatched tube and requisition labels.
Anatomical lesion site not specified.

Specimen Collection and Handling

Specimen Requirements

Test Requested	Required Requisition(s)	Specimen Type	Minimum Volume	Collection Kit
Syphilis PCR	General Test Requisition	Swab of lesion or lesion exudate in UTM	3 mL	Syphilis PCR (order# 390047)

Submission and Collection Notes

ALL sections of the General Test Requisition must be completed by an authorized health care provider for each specimen submitted. The site of the lesion must be specified in the Specimen Information section of the requisition.

For clinical specimens, label the specimen container(s) with the patient’s first and last name, collection date, and one other unique identifier such as the patient’s date of birth or Health Card Number. For additional information, see: Criteria for Acceptance of Patient Specimens. Failure to provide this information may result in rejection or testing delay.

Refer to the Syphilis PCR Lesion Collection for detailed collection instructions.

Limitations

PHO’s syphilis PCR LDT does not distinguish between the other subspecies of T. pallidum (T. pallidum subsp. endemicum (bejel), T. pallidum subsp. pertenue (yaws).

Storage and Transport

Place the specimen tube in the biohazard bag and seal. Then, place the completed General Test Requisition in the pouch at the front of the biohazard bag.

Following collection, store specimens at 2-8 °C and ship refrigerated to PHO. If transport is expected to be delayed for more than 48 hours, store specimens at -20°C and ship on dry ice.

All clinical specimens must be shipped in accordance to the Transportation of Dangerous Good Act.

Requisitions and Kit Ordering

Test Frequency and Turnaround Time (TAT)

Syphilis PCR is performed 2-3 times per week Monday to Friday. The turnaround time is up to 4 business days from specimen receipt at PHO.

Test Methods

Specimens are tested using a laboratory-developed (LDT) real-time PCR assay that detects T. pallidum DNA that has been validated by PHO for clinical use.

The syphilis PCR LDT includes two well established T. pallidum targets (tpp47, polA) and a human endogenous control target (RNaseP).

Interpretation

The detection of at least one of the two T. pallidum targets is considered positive. The following table provides possible test results and interpretation for syphilis PCR. Results should be interpreted in the context of the individual's presentation, clinical history, and other laboratory results, including syphilis serology.

Result	Interpretation	Notes
Detected	Treponema pallidum DNA detected.	If not already done, submit a serum specimen for syphilis serology.
Indeterminate	Detection of Treponema pallidum DNA is inconclusive.	An indeterminate result may be due to low bacterial quantity approaching the limit of detection of the assay or nonspecific reactivity in the specimen. Consider submitting a new specimen if clinically indicated. If not already done, submit a serum specimen for syphilis serology.
Not Detected	Treponema pallidum DNA not detected.	A negative result does not rule out syphilis infection. If not already done, submit a serum specimen for syphilis serology.
Invalid	The PCR test is invalid.	An invalid result may be due to inadequate specimen collection, extraction failure, or PCR inhibition. Submit a new specimen if clinically indicated. If not already done, submit a serum specimen for syphilis serology.

Reporting

Results are reported to the physician, authorized health care provider (General O. Reg 45/22, s.18) or submitter as indicated on the requisition.

Detected and indeterminate results are reported to the Medical Officer of Health as per the Ontario Health Protection and Promotion Act.